Biomedical subjects

Bradley R Cairns

Publications and source records attributed to Bradley R Cairns.

At least 19 recordsLinked to original sources

PubMed2026

BRD9 Degraders Unleash GBAF Chromatin Remodeling Activity in Synovial Sarcoma.

UNLABELLED: Synovial sarcoma incorporates the SS18::SSX fusion oncoprotein into GLTSCR1-containing BRG1/BRM and associated factors (GBAF) complexes, which confers a dependency on the GBAF subunit BRD9. However, synovial sarcoma clinical trials with multiple BRD9 degraders failed to achieve clinically impactful remissions. In this study, we identified a mechanistic framework to explain these results. BRD9 depletion served to blunt proliferation in synovial sarcoma harboring minimal genomic alterations, rare in trial participants. In cultured cells, xenografts, and recombinant-purified complexes, BRD9 loss did not affect GBAF assembly. Although BRD9 degradation in synovial sarcoma reduced GBAF enrichment at target loci, BRD9-less complexes maintained or increased chromatin accessibility and associated gene transcription. Biochemical assays with purified recombinant GBAF demonstrated increased nucleosome sliding in the absence of BRD9. Together, these findings show that BRD9 restrains GBAF activity, with BRD9 degradation increasing enzymatic remodeling and target gene expression by fusion oncoprotein-distributed GBAFs in synovial sarcoma. This subtle epigenetic disturbance creates a low hurdle for synovial sarcoma to surpass, limiting the therapeutic efficacy of BRD9 degraders. SIGNIFICANCE: BRD9 represses the GBAF chromatin remodeling complex, which causes enhanced rather than disrupted SS18::SSX complex activity following BRD9 degradation and explains the lack of efficacy of pharmacological BRD9 degraders in synovial sarcoma.

Sarcoma, Synovial↗

PubMed2026

SWI/SNF Alterations Define a Chromatin-Dependent Subtype of Urothelial Carcinoma.

PURPOSE: SWI/SNF (BAF) chromatin remodeling complex alterations are common in urothelial carcinoma, yet no biomarker-directed therapeutic strategies have been established for this population. We investigated whether BAF alterations delineate a biologically distinct, therapeutically actionable urothelial carcinoma subtype. EXPERIMENTAL DESIGN: We performed integrative genomic and transcriptomic analyses of 792 urothelial carcinoma tumors from the Oncology Research Information Exchange Network (ORIEN) and validated findings in the TCGA-BLCA cohort. Mechanistic studies incorporated RNA sequencing and ATAC-seq following histone deacetylase (HDAC) inhibition. Functional dependencies were assessed using patient-derived xenograft organoids and cell line models. Clinical relevance was explored in a biomarker-enriched investigator-initiated trial. RESULTS: Approximately half of urothelial carcinoma tumors exhibited BAF alterations, defining a previously unrecognized chromatin-altered molecular subtype characterized by activation of proliferative programs, loss of lineage identity, and altered metabolic signaling. This subtype was enriched for transcriptomic programs associated with HDAC inhibitor sensitivity and depleted of HDAC inhibitor resistance signatures. Mechanistically, HDAC inhibition induced widespread chromatin remodeling with reduced accessibility at AP-1 and TEAD-associated regions, and downregulation of E2F- and MYC-driven transcriptional networks. Functional studies confirmed enhanced HDAC inhibition sensitivity in ARID1A -mutated cell lines and a patient-derived organoid model. Early clinical observations demonstrated a durable responder treated with HDAC inhibitors and immunotherapy. CONCLUSIONS: BAF alterations define a chromatin-dependent tumor state in urothelial carcinoma that is selectively vulnerable to HDAC inhibition. Integrating genomic, epigenomic, functional, and early clinical evidence, these findings provide a rationale for biomarker-enriched clinical trials and HDAC inhibitor-based combination strategies in urothelial carcinoma.

Journal Article↗

PubMed2006

DNA translocation and loop formation mechanism of chromatin remodeling by SWI/SNF and RSC.

ATP-dependent chromatin-remodeling complexes (remodelers) modulate gene transcription by regulating the accessibility of highly packaged genomic DNA. However, the molecular mechanisms involved at the nucleosomal level in this process remain controversial. Here, we monitor the real-time activity of single ySWI/SNF or RSC complexes on single, stretched nucleosomal templates under tensions above 1 pN forces. We find that these remodelers can translocate along DNA at rates of approximately 13 bp/s and generate forces up to approximately 12 pN, producing DNA loops of a broad range of sizes (20-1200 bp, average approximately 100 bp) in a nucleosome-dependent manner. This nucleosome-specific activity differs significantly from that on bare DNA observed under low tensions and suggests a nucleosome-remodeling mechanism through intranucleosomal DNA loop formation. Such loop formation may provide a molecular basis for the biological functions of remodelers.

Adenosine Triphosphate↗

PubMed2006

Dephosphorylation and genome-wide association of Maf1 with Pol III-transcribed genes during repression.

Nutrient deprivation and various stress conditions repress RNA polymerase III (Pol III) transcription in S. cerevisiae. The signaling pathways that relay stress and nutrient conditions converge on the conserved protein Maf1, but how Maf1 integrates environmental conditions and couples them to transcriptional repression is largely unknown. Here, we demonstrate that Maf1 is phosphorylated in favorable conditions, whereas diverse unfavorable conditions lead to rapid Maf1 dephosphorylation, nuclear localization, physical association of dephosphorylated Maf1 with Pol III, and Maf1 targeting to Pol III-transcribed genes genome wide. Furthermore, Maf1 mutants defective in full dephosphorylation display maf1Delta phenotypes and are compromised for both nuclear localization and Pol III association. Repression conditions also promote TFIIIB-TFIIIC interactions in crosslinked chromatin. Taken together, Maf1 appears to integrate environmental conditions and signaling pathways through its phosphorylation state, with stress leading to dephosphorylation, association with Pol III at target loci, alterations in basal factor interactions, and transcriptional repression.

Amino Acid Sequence↗

PubMed2006

ING2 PHD domain links histone H3 lysine 4 methylation to active gene repression.

Dynamic regulation of diverse nuclear processes is intimately linked to covalent modifications of chromatin. Much attention has focused on methylation at lysine 4 of histone H3 (H3K4), owing to its association with euchromatic genomic regions. H3K4 can be mono-, di- or tri-methylated. Trimethylated H3K4 (H3K4me3) is preferentially detected at active genes, and is proposed to promote gene expression through recognition by transcription-activating effector molecules. Here we identify a novel class of methylated H3K4 effector domains--the PHD domains of the ING (for inhibitor of growth) family of tumour suppressor proteins. The ING PHD domains are specific and highly robust binding modules for H3K4me3 and H3K4me2. ING2, a native subunit of a repressive mSin3a-HDAC1 histone deacetylase complex, binds with high affinity to the trimethylated species. In response to DNA damage, recognition of H3K4me3 by the ING2 PHD domain stabilizes the mSin3a-HDAC1 complex at the promoters of proliferation genes. This pathway constitutes a new mechanism by which H3K4me3 functions in active gene repression. Furthermore, ING2 modulates cellular responses to genotoxic insults, and these functions are critically dependent on ING2 interaction with H3K4me3. Together, our findings establish a pivotal role for trimethylation of H3K4 in gene repression and, potentially, tumour suppressor mechanisms.

Amino Acid Motifs↗

PubMed2006

Structure and function of the SWIRM domain, a conserved protein module found in chromatin regulatory complexes.

The SWIRM domain is a module found in the Swi3 and Rsc8 subunits of SWI/SNF-family chromatin remodeling complexes, and the Ada2 and BHC110/LSD1 subunits of chromatin modification complexes. Here we report the high-resolution crystal structure of the SWIRM domain from Swi3 and characterize the in vitro and in vivo function of the SWIRM domains from Saccharomyces cerevisiae Swi3 and Rsc8. The Swi3 SWIRM forms a four-helix bundle containing a pseudo 2-fold axis and a helix-turn-helix motif commonly found in DNA-binding proteins. We show that the Swi3 SWIRM binds free DNA and mononucleosomes with high and comparable affinity and that a subset of Swi3 substitution mutants that display growth defects in vivo also show impaired DNA-binding activity in vitro, consistent with a nucleosome targeting function of this domain. Genetic and biochemical studies also reveal that the Rsc8 and Swi3 SWIRM domains are essential for the proper assembly and in vivo functions of their respective complexes. Together, these studies identify the SWIRM domain as an essential multifunctional module for the regulation of gene expression.

Amino Acid Sequence↗

PubMed2006

Mechanisms for nucleosome movement by ATP-dependent chromatin remodeling complexes.

Chromatin remodeling complexes (remodelers) are a set of diverse multi-protein machines that reposition and restructure nucleosomes. Remodelers are specialized, containing unique proteins that assist in targeting, interaction with modified nucleosomes, and performing specific chromatin tasks. However, all remodelers contain an ATPase domain that is highly similar to known DNA translocases/helicases, suggesting that DNA translocation is a property common to all remodelers. Here we examine the different reactions they perform in vitro, focusing on the SWI/SNF and the ISWI complexes, and explore how DNA translocation might be utilized to execute various remodeling processes.

Adenosine Triphosphatases↗

PubMed2006

Chromatin remodelling: the industrial revolution of DNA around histones.

Chromatin remodellers are specialized multi-protein machines that enable access to nucleosomal DNA by altering the structure, composition and positioning of nucleosomes. All remodellers have a catalytic ATPase subunit that is similar to known DNA-translocating motor proteins, suggesting DNA translocation as a unifying aspect of their mechanism. Here, we explore the diversity and specialization of chromatin remodellers, discuss how nucleosome modifications regulate remodeller activity and consider a model for the exposure of nucleosomal DNA that involves the use of directional DNA translocation to pump 'DNA waves' around the nucleosome.

Animals↗

PubMed2006

Zebra fish Dnmt1 and Suv39h1 regulate organ-specific terminal differentiation during development.

DNA methylation and histone methylation are two key epigenetic modifications that help govern heterochromatin dynamics. The roles for these chromatin-modifying activities in directing tissue-specific development remain largely unknown. To address this issue, we examined the roles of DNA methyltransferase 1 (Dnmt1) and the H3K9 histone methyltransferase Suv39h1 in zebra fish development. Knockdown of Dnmt1 in zebra fish embryos caused defects in terminal differentiation of the intestine, exocrine pancreas, and retina. Interestingly, not all tissues required Dnmt1, as differentiation of the liver and endocrine pancreas appeared normal. Proper differentiation depended on Dnmt1 catalytic activity, as Dnmt1 morphants could be rescued by active zebra fish or human DNMT1 but not by catalytically inactive derivatives. Dnmt1 morphants exhibited dramatic reductions of both genomic cytosine methylation and genome-wide H3K9 trimethyl levels, leading us to investigate the overlap of in vivo functions of Dnmt1 and Suv39h1. Embryos lacking Suv39h1 had organ-specific terminal differentiation defects that produced largely phenocopies of Dnmt1 morphants but retained wild-type levels of DNA methylation. Remarkably, suv39h1 overexpression rescued markers of terminal differentiation in Dnmt1 morphants. Our results suggest that Dnmt1 activity helps direct histone methylation by Suv39h1 and that, together, Dnmt1 and Suv39h1 help guide the terminal differentiation of particular tissues.

Animals↗

PubMed2006

Rsc4 connects the chromatin remodeler RSC to RNA polymerases.

RSC is an essential, multisubunit chromatin remodeling complex. We show here that the Rsc4 subunit of RSC interacted via its C terminus with Rpb5, a conserved subunit shared by all three nuclear RNA polymerases (Pol). Furthermore, the RSC complex coimmunoprecipitated with all three RNA polymerases. Mutations in the C terminus of Rsc4 conferred a thermosensitive phenotype and the loss of interaction with Rpb5. Certain thermosensitive rpb5 mutations were lethal in combination with an rsc4 mutation, supporting the physiological significance of the interaction. Pol II transcription of ca. 12% of the yeast genome was increased or decreased twofold or more in a rsc4 C-terminal mutant. The transcription of the Pol III-transcribed genes SNR6 and RPR1 was also reduced, in agreement with the observed localization of RSC near many class III genes. Rsc4 C-terminal mutations did not alter the stability or assembly of the RSC complex, suggesting an impact on Rsc4 function. Strikingly, a C-terminal mutation of Rsc4 did not impair RSC recruitment to the RSC-responsive genes DUT1 and SMX3 but rather changed the chromatin accessibility of DNases to their promoter regions, suggesting that the altered transcription of DUT1 and SMX3 was the consequence of altered chromatin remodeling.

Chromatin↗

PubMed2005

Genome-wide dynamics of Htz1, a histone H2A variant that poises repressed/basal promoters for activation through histone loss.

Histone variants help specialize chromatin regions; however, their impact on transcriptional regulation is largely unknown. Here, we determined the genome-wide localization and dynamics of Htz1, the yeast histone H2A variant. Htz1 localizes to hundreds of repressed/basal Pol II promoters and prefers TATA-less promoters. Specific Htz1 deposition requires the SWR1 complex, which largely colocalizes with Htz1. Htz1 occupancy correlates with particular histone modifications, and Htz1 deposition is partially reliant on Gcn5 (a histone acetyltransferase) and Bdf1, an SWR1 complex member that binds acetylated histones. Changes in growth conditions cause a striking redistribution of Htz1 from activated to repressed/basal promoters. Furthermore, Htz1 promotes full gene activation but does not generally impact repression. Importantly, Htz1 releases from purified chromatin in vitro under conditions where H2A and H3 remain associated. We suggest that Htz1-bearing nucleosomes are deposited at repressed/basal promoters but facilitate activation through their susceptibility to loss, thereby helping to expose promoter DNA.

Acetylation↗

PubMed2005

The RSC chromatin remodeling complex bears an essential fungal-specific protein module with broad functional roles.

RSC is an essential and abundant ATP-dependent chromatin remodeling complex from Saccharomyces cerevisiae. Here we show that the RSC components Rsc7/Npl6 and Rsc14/Ldb7 interact physically and/or functionally with Rsc3, Rsc30, and Htl1 to form a module important for a broad range of RSC functions. A strain lacking Rsc7 fails to properly assemble RSC, which confers sensitivity to temperature and to agents that cause DNA damage, microtubule depolymerization, or cell wall stress (likely via transcriptional misregulation). Cells lacking Rsc14 display sensitivity to cell wall stress and are deficient in the assembly of Rsc3 and Rsc30. Interestingly, certain rsc7delta and rsc14delta phenotypes are suppressed by an increased dosage of Rsc3, an essential RSC member with roles in cell wall integrity and spindle checkpoint pathways. Thus, Rsc7 and Rsc14 have different roles in the module as well as sharing physical and functional connections to Rsc3. Using a genetic array of nonessential null mutations (SGA) we identified mutations that are sick/lethal in combination with the rsc7delta mutation, which revealed connections to a surprisingly large number of chromatin remodeling complexes and cellular processes. Taken together, we define a protein module on the RSC complex with links to a broad spectrum of cellular functions.

Chromatin Assembly and Disassembly↗

PubMed2005

Chromatin remodeling through directional DNA translocation from an internal nucleosomal site.

The RSC chromatin remodeler contains Sth1, an ATP-dependent DNA translocase. On DNA substrates, RSC/Sth1 tracks along one strand of the duplex with a 3' --> 5' polarity and a tracking requirement of one base, properties that may enable directional DNA translocation on nucleosomes. The binding of RSC or Sth1 elicits a DNase I-hypersensitive site approximately two DNA turns from the nucleosomal dyad, and the binding of Sth1 requires intact DNA at this location. Results with various nucleosome substrates suggest that RSC/Sth1 remains at a fixed position on the histone octamer and that Sth1 conducts directional DNA translocation from a location about two turns from the nucleosomal dyad, drawing in DNA from one side of the nucleosome and pumping it toward the other. These studies suggest that nucleosome mobilization involves directional DNA translocation initiating from a fixed internal site on the nucleosome.

Animals↗

PubMed2005

Distinct roles for the RSC and Swi/Snf ATP-dependent chromatin remodelers in DNA double-strand break repair.

The failure of cells to repair damaged DNA can result in genomic instability and cancer. To efficiently repair chromosomal DNA lesions, the repair machinery must gain access to the damaged DNA in the context of chromatin. Here we report that both the RSC and Swi/Snf ATP-dependent chromatin-remodeling complexes play key roles in double-strand break (DSB) repair, specifically by homologous recombination (HR). RSC and Swi/Snf are each recruited to an in vivo DSB site but with distinct kinetics. We show that Swi/Snf is required earlier, at or preceding the strand invasion step of HR, while RSC is required following synapsis for completion of the recombinational repair event.

Adenosine Triphosphate↗

PubMed2005

Histone trimethylation by Set1 is coordinated by the RRM, autoinhibitory, and catalytic domains.

Trimethylation of lysine 4 of histone H3 occurs at the 5' end of active genes and is catalyzed by Set1 in Saccharomyces cerevisiae. Trimethylation requires histone H2B ubiquitylation and the PAF1 complex, which are linked to transcription elongation, but how they activate Set1 is not known. Set1 also bears several conserved domains with uncharacterized contributions to activity. Here, we isolated dominant hyperactive SET1(D) alleles, which revealed a complex interplay among Set1 regulatory domains. Remarkably, the RNA-recognition motif (RRM) of Set1 is required for H3K4 trimethylation, but not dimethylation. Also, a central autoinhibitory domain was identified that opposes RRM function by inhibiting trimethylation. Furthermore, a G990E replacement in the catalytic domain conferred Set1 hyperactivity and restored trimethylation to a Set1 derivative bearing mutations in the RRM domain. Surprisingly, certain SET1(D) alleles also partially restored trimethylation to strains lacking histone H2B ubiquitylation or Paf1. Taken together, our data suggest that the catalytic domain of Set1 integrates opposing inputs from the RRM and autoinhibitory domains to link properly H3K4 methylation to the transcript elongation process.

Alleles↗

PubMed2005

Chromatin remodeling complexes: strength in diversity, precision through specialization.

Chromatin remodeling complexes all have an ATPase subunit important for nucleosome remodeling, but are diversified and specialized by additional associated proteins. Remodelers have well-established roles in a wide range of chromosomal processes, including transcriptional regulation and chromatin assembly. Recent work, however, has revealed remarkable new functions for remodelers, such as histone variant deposition, cohesin function, and RNA transcript elongation and termination. Remodeler complexes are tailored both compositionally and mechanistically to perform particular chromatin functions.

Chromatin↗

PubMed2004

Around the world of DNA damage INO80 days.

DNA damage is detected and repaired in the context of chromatin. In this issue of Cell, van Attikum et al. (2004) and Morrison et al. (2004) demonstrate that double-stranded break repair involves the recruitment of a specialized chromatin remodeling complex, INO80, through an interaction with phosphorylated histone H2A.

Adenosine Triphosphate↗

PubMed2004

Tandem bromodomains in the chromatin remodeler RSC recognize acetylated histone H3 Lys14.

The coordination of chromatin remodeling with chromatin modification is a central topic in gene regulation. The yeast chromatin remodeling complex RSC bears multiple bromodomains, motifs for acetyl-lysine and histone tail interaction. Here, we identify and characterize Rsc4 and show that it bears tandem essential bromodomains. Conditional rsc4 bromodomain mutations were isolated, and were lethal in combination with gcn5Delta, whereas combinations with esa1 grew well. Replacements involving Lys14 of histone H3 (the main target of Gcn5), but not other H3 or H4 lysine residues, also conferred severe growth defects to rsc4 mutant strains. Importantly, wild-type Rsc4 bound an H3 tail peptide acetylated at Lys14, whereas a bromodomain mutant derivative did not. Loss of particular histone deacetylases suppressed rsc4 bromodomain mutations, suggesting that Rsc4 promotes gene activation. Furthermore, rsc4 mutants displayed defects in the activation of genes involved in nicotinic acid biosynthesis, cell wall integrity, and other pathways. Taken together, Rsc4 bears essential tandem bromodomains that rely on H3 Lys14 acetylation to assist RSC complex for gene activation.

Acetylation↗